Hexagrammos otakiiis an important kind of economic fish species in East Asia. However, wild H. otakii resources have declined sharply in recent years as a result of human disturbance and habitat destruction. Thus, it is crucial to protect the current resources of H. otakii. In this study, 35 novel single nucleotide polymorphism (SNP) markers were developed based on restriction-site associated DNA sequencing. The results showed that the observed heterozygosity and expected heterozygosity ranged from 0.1875 to 0.6562 and 0.2679 to 0.5079, respectively. The minor allele frequency ranged from 0.1875 to 0.4375. Polymorphic information content ranged from 0.229 to 0.375. Six SNPs were found to be deviated significantly from the HWE (P < 0.05). These SNP markers will serve as a useful tool for genetic studies and population evaluation aimed at the conservation of H. otakii.
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